Tumour necrosis factor alpha (TNF-α) and tumour necrosis factor beta (TNF-β) exhibit a significant overlap in function. This homology has resulted in these cytokines being collectively referred to as tumour necrosis factor (TNF). TNF exhibits a powerful pleiotropic role in mediating the proinflammatory immune response and has also been shown to have involvement in control of cell proliferation, differentiation and apoptosis. TNF mediates these effects by binding to the type I and II TNF receptors (TNFR), which are cell surface receptors that are expressed on TNF responsive cells.
The use of TNF antagonists, such as anti-TNF antibodies, soluble TNF receptor proteins and TNF receptor-Fc fusion proteins has shown that the proinflammatory effects mediated by TNF, such as induction of the expression of proinflammatory cytokine expression (IL-1, IL-8) and tissue destruction, can be reversed. Furthermore, the use of recombinant TNFR-Fc fusion proteins such as ENBREL (Etanercept, Immunex) has shown that TNF antagonism can be used to treat TNF-associated conditions, in particular rheumatoid arthritis.
Accordingly, the inhibition of TNF using tumour necrosis factor receptor (TNFR)-immunoglobulin Fc domain fusion proteins or anti-TNF neutralising monoclonal antibodies has been proven to be a successful therapeutic approach for the treatment of a variety of human inflammatory diseases, including rheumatoid arthritis and psoriatic arthritis.
Companion animals, such as dogs, develop inflammatory diseases similar to those which occur in humans, with examples being rheumatoid arthritis (RA), osteoarthritis, immune-mediated polyarthritidies, plasmatic-lymphocytic synovitis, systemic lupus erythematosis (SLE), vasculitis and a variety of autoimmune skin diseases. It is estimated that one in five adult dogs in the USA has arthritis and dogs have been used as models of human joint disease, e.g. for osteoarthritis, anterior cruciate ligament disruption and meniscal damage.
The role of TNF in the occurrence of inflammation in dogs has been extensively documented. For example, it has been observed that there is increased secretion of TNF alpha in cell infiltrates of synovial fluid of dogs presenting with stifle arthritis TNF and the type II TNF receptor has been shown to be significantly elevated in the central and peripheral retina of dogs with glaucoma as part of a broad inflammatory response. Treatment of dogs with the human TNFR-Fc fusion protein Etanercept ((huTNFR-Fc) Immunex, a TNF antagonist) reduced myocardial injury by approximately 25-40% following ischaemia-reperfusion induced by balloon occlusion in a closed chest model of human heart disease, with a concomitant reduction in associated inflammatory markers such as ICAM-1 and NF-kB. Similarly, a 60% reduction in infarct size in an open chest dog model of ischaemia reperfusion using 2 mg/kg TNFR-Fc has also been demonstrated. However, the use of huTNFR-Fc as a therapeutic agent for the treatment of diseases of the dog is not indicated beyond this modelling of human disease due to the immunogenicity of human proteins when injected into dogs. Furthermore, the IgG Fc domain of Etanercept is complement recruiting and hence undesirable in the context of an inflammatory disease, due to the immune response which is mediated.
Anti-canine TNF monoclonal antibodies have been used to detect low levels of TNF by capture ELISA in supernatants of canine PBMCs treated with lipopolysaccharide (LPS) and TNF alpha expression has also been reported in skin samples of canine hemangiopericytoma, tricoblastoma, lipoma and mastocytoma. TNF-alpha and TNF receptors are present in canine articular cartilage in an induced model of osteoarthritis, while Adalimumab, a humanised monoclonal antibody to TNF, has been tested in two dogs with exfoliative cutaneous lupus erythematosus (ECLE) (0.5 mg/kg every 2 weeks for 8 weeks), but disease progression was shown to be unaltered, with serum TNF-alpha levels remaining unchanged. Immunogenicity to Adalimumab would potentially have caused the lack of efficacy in these repeat dosing studies. In particular, neutralising antibodies which may be raised against Adalimumab would prevent repeat dosing and therefore restrict the longer term effectiveness of such a therapeutic approach.
Accordingly, due to the involvement of TNF in a wide range of inflammatory mediated conditions in canines, there is a need for inhibitors of canine TNF that can be used for the long-term inhibition of TNF in order to treat TNF-associated disorders and inflammatory conditions in dogs.